The Section of Neuropathology supports the primary goal of the CBDB which uses evidence of genetic association to generate and test hypotheses about how specific genes affect brain development, plasticity and function and how these effects relate to biology leading to potential treatment of illness. Our core lab functions as a component of the gene-based hypothesis testing and biologic validation program in the Branch. We have been successful in translating clinical associations into molecular mechanisms. Over the past year, our analysis has shown alternative splicing in postmortem human brain tissue and the mechanism of genetic association (e.g. Disrupted-In-Schizophrenia 1 (DISC1), Neuregulin 1 (NRG1), potassium voltage-gated channel (KCNH2)) appear to relate specifically to regulation of expression of novel splice forms. We have noted on previous reports that many of the schizophrenia associated splice variants are preferentially expressed in fetal brain, implicating early brain development as a critical time for the molecular effects of risk associated genetic variation. We continue to develop expression profile datasets using several commercial platforms and high density chip-based SNP genotyping to identify common variants associated with gene expression. Through the use of highly technical and sensitive instruments we have validated genes showing association with schizophrenia, related intermediate phenotypes, and identified potentially critical genetic markers for other neuropsychiatric disorders. Statistical associations of complex genetic disorders typically show small effects, and identifying functional mutations is very complicated, so our goal is to validate genetic associations with convergent biological data that specific genetic variations affect aspects of brain function and molecular biology. Our on-going research is based on the assumption that schizophrenia is a combination of traits that interact with each other, other genes and the environment to produce the complex clinical phenotype. We hypothesize that elucidating mechanisms of disease will help us to identify novel therapeutic targets from association of genetic variation with relevant biologic intermediate phenotypes. Our research this year has strengthened the validity of this approach as evidenced in our studies of KCNH2 (Huffaker, 2009) and DISC1 (Nakata, Lipska, 2010). Last year, we reported finding a novel isoform of a candidate schizophrenia susceptibility gene, KCNH2, which modulates organized neuronal firing. This activity is critical for cortical information processing which is notably deficient in schizophrenia. We identified a novel primate-specific 3.1 isoform found only in brain. We found isoform 3.1 mRNA levels to be comparable to KCNH2 in prefrontal cortex and hippocampus but over 1000 fold lower in heart. Postmortem expression analysis revealed a 2.5 fold increase in isoform 3.1 in schizophrenic hippocampus relative to normals. There were also clinical associations in the same genetic and allelic variations in the CBDB family dataset and a German case control collaborative dataset. A meta-analysis of 5 independent clinical samples involving over 3500 subjects showed statistically significant association of single nucleotide polymorphisms (SNPs) in KCNH2 with schizophrenia. Risk-associated SNPs also predicted: 1) increased isoform 3.1 mRNA expression in postmortem human hippocampus, 2) lower IQ scores and speed of cognitive processing in healthy controls, 3) altered hippocampal structure and functional magnetic resonance imaging (fMRI) physiological signals in the hippocampus and prefrontal cortex in healthy subjects. Electrophysiological characterization in primary cortical neurons revealed that overexpression of Isoform 3.1 resulted in a rapidly deactivating potassium current and a high-frequency, non-adapting firing pattern. These findings were a collaborative effort and we will pursue modifying the effect of this novel KCNH2 3.1 isoform on neuronal function as a new therapeutic target. This year, we delved further into our study of DISC1. DISC1 has been identified as one of the most promising schizophrenia-susceptibility genes. A large number of DISC1 alternatively spliced variants were identified in human postmortem brain but the molecular mechanism of association remains unclear. These include novel transcripts that lack exon 3 (&#8710;3), exons 7 and 8 (&#8710;7&#8710;8), an exon 3 insertion variant (extra short variant-1, (Esv1)), and a group of transcripts resulting from intergenic splicing between TSNAX (Translin-associated factor X) and DISC1. We also report that the group of short isoforms &#8710;3, &#8710;7&#8710;8, and Esv1 encoded truncated DISC1 proteins that were more abundantly expressed during fetal development than during postnatal ages, and that their expression in hippocampus is significantly higher in patients with schizophrenia. In addition, two DISC1 coding SNPs, which may be associated with risk for schizophrenia, showed significant effects on expression of isoforms &#8710;3 and &#8710;7&#8710;8. It is interesting that the same allele that predicts higher expression levels in hippocampus is also associated with higher expression of DISC1 &#8710;7&#8710;8 in lymphoblasts. The allelic heterogeneity associated with DISC1 risk for schizophrenia could be the result of different SNPs impacting different alternative transcripts in so far as the novel isoform lacks the C terminus, they may not be able to interact with a number of DISC1 binding partners. We have previously reported that expression of several DISC1 binding partners are reduced in the hippocampus of patients. We will continue our exploration of a molecular mechanism of a DISC1 genetic risk association via alternative splicing processes.